Soil-transmitted helminths (STHs) are common nematode parasites of the human intestinal tract. The main STHs are Ascaris, hookworms, and whipworms. These parasites infect > 400,000,000 children worldwide, leading to significant malnutrition, physical stunting, cognitive deficiencies, school absenteeism, lower future earnings, lower educational status, vaccine failure, and immune deficiencies. The parasites also complicate pregnancies, cause lethargy, and result in low worker productivity in adults. Recent estimates have placed the burden of STH disease comparable of that malaria and cancer. In all these ways, the devastating impact of STH infections affect childhood health and development, maternal and adult health, and economic productivity in such a way as to trap endemic communities in a cycle of poverty, underdevelopment, and disease. Currently only one drug, albendazole, has adequate efficacy for single-dose mass drug administration. Albendazole has moderate and highly variable efficacy against hookworms and very poor efficacy against whipworms. Recent data have revealed instances of unusually low or reduced efficacy of albendazole over time against hookworms and Ascaris. New and improved classes of anthelmintics to treat STHs are urgently needed. This project proposes a radical new approach for deworming-use of the vertebrate-safe Bacillus thuringiensis crystal (Cry) protein, Cry5B (and potentially Cry14A), delivered via potent and unique formulations or via an engineered probiotic bacterium, Bacillus subtilis natto. Cry proteins are generally regarded as safe to humans. When given orally, single-dose Cry5B can cure hookworm infections in hamsters and Ascaris infections in pigs; whipworms are sensitive to Cry5B in vitro. Cry5B efficacy can be boosted using biological encapsulations. Probiotic Bacillus subtilis natto, eaten for centuries by the Japanese, can be engineered to express bioactive Cry5B with strong anthelmintic properties. The aims of this project are (Aim 1) to optimize biological formulation of Cry5B protein and expression of Cry5B in probiotic B. subtilis natto, testing in vitro activity against all the parastes and in vivo activity against hookworms and whipworms in rodents and Ascaris in pigs; (Aim 2) develop and test against all parasites the next generation Cry protein anthelmintic with promising properties, Cry14A; and (Aim 3) investigate potential powerful synergistic drug combinations with Cry5B to delay or prevent the evolution of parasite resistance. At the conclusion of this study, we will select a delivery form of Cry5B to be advanced into pre-clinical and future clinical trials based upon 1) highest efficacy against parasites in vivo, 2) lowest delivery cost, and 3) discussions with regulators (FDA).